ATP-sensitive potassium (K-ATP) channels, coupling cell metabolism to cell membrane potential, are involved in brain diseases including stroke. Emerging evidence shows that astrocytes play important roles in the pathophysiology of cerebral ischemia. Kir6.1, a pore-forming subunit of K-ATP channel, is prominently expressed in astrocytes and participates in regulating its function. However, the exact role of astrocytic Kir6.1-containg K-ATP channel (Kir6.1/K-ATP) in ischemic stroke remains unclear. Here, we found that astrocytic Kir6.1 knockout (KO) mice exhibited larger infarct areas and more severe brain edema and neurological deficits in middle cerebral artery occlusion stroke model. Both activated gliosis and neuronal loss were aggravated in astrocytic Kir6.1 KO mice. Furthermore, the protein levels of pro-apoptotic protein Bcl-2 associated X (Bax) and active caspase-3 were up-regulated and the expression of anti-apoptotic protein Bcl-2 was down-regulated in astrocytic Kir6.1 KO mice. This is accompanied by enhanced endoplasmic reticulum stress (ER stress) responses in brain tissues and in astrocytes during ischemia/reperfusion (I/R) injury. Finally, inhibition of ER stress rescued astrocyte apoptosis induced by Kir6.1 deletion during I/R injury. Collectively, our findings reveal that astrocytic Kir6.1/K-ATP channel protects brain from cerebral ischemia/reperfusion injury through inhibiting ER stress and suggest that astrocytic Kir6.1/K-ATP channel is a promising therapeutic target for ischemic stroke.